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標題: 利用轉錄體資料分析並預測人類微核糖核酸之轉錄起始點
Identification of human microRNA transcriptional start sites from transcriptome
作者: 許郁彬
Hsu, Yu-Pin
Contributors: 謝立青
Li-Ching Hsieh
基因體暨生物資訊學研究所
關鍵字: 微核糖核酸;轉錄起始位;轉錄因子結合位;上游調控
MicroRNA;Transcription Start Site;TSS;Transcription Factor Binding Site;TFBS;upstream regulation
日期: 2013
Issue Date: 2013-11-19 12:01:57 (UTC+8)
Publisher: 基因體暨生物資訊學研究所
摘要: 過去許多研究基因調控的研究者試圖藉由分析轉錄起始點 (transcriptional start site;TSS)來標定目標基因的序列起始位置及調控區域。由於次世代定序技術的發展使得分析預測轉錄起始點的方法也越來越多樣化,如利用ChIP-seq (Chromatin immunoprecipitation sequencing)技術來分析RNA PolⅡ的接合位置來獲取轉錄起始點資訊是最基本的方法之一,同時也搭配分析訊息核糖核酸 (mRNA)的5’端序列 (5’-end SAGE[1])、5’端CAP的序列 (CAGE[2]),或是組蛋白 (histone)修飾位置來協助找尋轉錄起始點。在過去大多數研究者僅採用其中一種或是兩種方法來找尋轉錄起始點,另外有一些研究者結合其他非傳統ChIP-seq科技來定位轉錄起始點,如利用DNAase-seq [3] (DNase I hypersensitive sites sequencing) 及FAIRE-seq [4] (Formaldehyde-Assisted Isolation of Regulatory Elements sequencing) 技術並搭配序列保守性或是機器學習 [5]的方式。這些方法也被用來分析 microRNA 的轉錄起始點。我們則嘗試利用RNA-seq 資料(即Transciptome的定序資料)來尋找人類miRNA之轉錄起始點,我們在轉錄起始點的位置附近看到獨特的表現量變化,解決splice junction的定位問題後,我們嘗試組合完整的pri-miRNA,提供更完整的pri-miRNA資訊,獨立分析transcriptome資料不僅僅能夠預測miRNA 轉錄起始點位置,也可以針對不同類型的轉錄起始點進行分析 [6],同時也提供其他研究者不同的特徵選擇。統合不同的實驗數據來分析預測及定義轉錄起始點,對於研究基因調控的研究者而言更具有幫助。
關鍵字:微核糖核酸、轉錄起始位、轉錄因子結合位、上游調控
Identifying gene transcription start sites (TSSs) is the basis for determining gene upstream regulatory region. With the progress of the next generation sequencing (NGS) technologies, many kinds of NGS data have been utilized to identify TSSs. For example, RNA PolⅡChIP-seq (Chromatin immunoprecipitation sequencing) is used to recognize RNA PolⅡbinding sites and therefore is a direct way to identify TSSs. In addition, some other kinds of data like nucleosome-seq, DNase-seq (DNase I hypersensitive sites sequencing) and FAIRE-seq (Formaldehyde-Assisted Isolation of Regulatory Elements sequencing), which are associated with histone modification sites, hyper-sensitive sites and regulatory activity, respectively, are alone or combined to determine TSSs. Several statistical or machine learning methods have been developed for finding and characterizing TSSs. These datasets and methods can be also used to locate miRNA TSSs because it is believed that most miRNA genes, like protein coding genes, require RNA Pol II for expression.
In this thesis, we utilized RNA-seq data, i.e. transcriptome sequencing data, to locate miRNA TSSs. We observed that there are signals of RNA-seq sequences near the TSS of well-annotated protein coding genes so we proposed that the miRNA TSSs may also have similar pattern. We then located splice junction region and tried to complete primary transcripts of miRNAs (i.e. pri-miRNAs) and then miRNA TSSs were identified. This approach can not only locate miRNA TSSs but also give information to characterize TSSs of other gene types. Information obtained from transcriptome sequencing data could be a new feature for predicting gene TSSs and analyzing gene regulation.
KEY WORDS
MicroRNA, Transcription Start Site, TSS, Transcription Factor Binding Site, TFBS, upstream regulation
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